This invention relates to an improved antigenic substance for testing for tuberculosis and to the method for preparing such antigenic substance.
It has been known for many years that exposure of a human being to the disease commonly known as tuberculosis, resulting from an infection by Mycobacterium tuberculosis (M. tb.) may be tested for by an intradermal injection of antigenic material derived from tubercle bacilli or their products. The antigenic material, tuberculin, induces sensitized lymphocytes to initiate a delayed hypersensitivity response at the site of the injection. The subsequent swelling at that site is known as edema or induration, and the reddening in the area is known as erythema. After a selected lapse of time, generally 48 to 72 hours, the size of the edematous swelling is measured and compared to previously established statistical standards that relate size of reaction after a particular dose of antigen to probability and recentness of prior infection with M. tb. Thus, an evaluation is made as to whether the injectee is a "positive" or "negative" reactor.
A number of antigenic materials or tuberculins exist or have been suggested, and a number of intradermal injectors have been used. The classical standard of injector is a needle and syringe using a technique known throughout medical literature as the "Mantoux test", a technique that requires considerable skill both in administration and in evaluating the results. Other injectors and techniques include and have included a simple scratch test, the "Vollmer" patch test, multiple scratch scarifications by an instrument known as the Heaf gun (also known by the Trademark "Sterneedle"), or by simultaneous skin puncture and injection as effected by instruments known as "Mono-Vacc" of Lincoln Laboratories and "Tine Test" of Lederle Laboratories. For reasons that are unimportant here, the Mantoux test is generally accepted as a standard diagnostic test while use of the other instruments is generally considered a "screening" test useful as a first test in screening from a large group of humans being tested, those who appear to be clearly positive reactors. Some of the test noted, such as the Vollmer patch test, have been discarded as unreliable.
The problem of "unreliability" is constantly present as a factor even with use of the Mantoux test. A major reason for the unreliability factor stems from the nature of the antigenic material used by the test instruments and test techniques. Until now the two tuberculins used are known in the medical literature as "Old Tuberculin" (OT) and "Purified Protein Derivative" (PPD), a derivative of OT.
The medical literature has for years noted the existence of the unreliability factor under the term "false positives" with respect to use of OT and PPD. These undesirable reactions stem principally from stimulation of lymphocytes previously sensitized by prior exposure or infection by other bacteria, such as "atypical mycobacteria" which are closely related to M. tb. Certain antigens are shared in common by M. tb. and the other mycobacteria. If these shared or "cross-reactive antigens" are present in the tuberculin injected into the skin, an individual previously infected by mycobacteria other than M. tb. could give a positive response.
The existence of false positives resulting from use of OT or PPD with all types of injectors or scratch instruments has created a real problem of long standing duration to the medical community. It has been widely acknowledged that a specific antigenic material, that may be used in a reliable and efficient manner as a diagnostic test for M. tb., is greatly needed.
It has been a postulate of some researchers in the field that isolation of a single antigen specific to M. tb. would provide the ideal antigenic material for diagnostic skin testing for M. tb., and for diagnostic tests of M. tb. sensitization or infection, in vitro.
Many studies have been made of the antigenic "mix" of M. tb. culture filtrates toward the end of somehow selecting and isolating an antigen of diagnostic reliability and great specificity; many techniques have been devised for such mycobacterial antigen isolation. See, for example, American Review of Respiratory Diseases, Vol. 89, No. 1, January, 1964 P. 29 et seq. and Vol. 92, December, Part 2, 1965, P. 19 et seq. However, until now, it appears that the key to preparation of an antigenic material that has both high specificity and high sensitivity has eluded the search by investigators, despite the fact that antigens specific for M. tb. appear to exist and the literature describing preparative and analytical methods for the separation and isolation of antigens of M. tb. has been available for many years.